Many diseases are today curable only by a transplantation of tissue or an organ, such as a kidney or heart. It is sometimes possible to locate a living donor with immunological markers compatible with the transplant recipient, although organ donation by a living donor involves great risks and possible deleterious health effects for the donor. Without any available living donor, the organ must be obtained from a heartbeating human cadaver of high quality and, again, there must be a good immunological match between the donor and the recipient. The situation today is a steadily increasing demand for human organs suitable for transplantation and the gap between said demand and the availability of organs is likely to grow even wider in view of the continuing improvements made in transplantation procedures and outcome. The most promising possible answer to this problem is xenotransplantation, i.e. transplantation of tissue or organs between different species. For human patients, the pig is considered the most suitable donor species for medical, practical, ethical and economical reasons.
The main problem in xenografting between discordant species, such as pig to human, is the hyperacute rejection (HAR), which leads to a cessation of the blood flow within minutes following a transplantation. Even though other mechanisms of rejection will ensue after HAR, the general belief is that if HAR could be prevented, the patient's immune system may undergo a process of accommodation, whereafter a conventional immunosuppressive regimen could maintain the compatibility of the patient and the xenograft.
The HAR is caused by preformed, natural antibodies in the receiving species reacting with antigens on the endothelium in donor organs, an interaction which leads to complement and endothelial cell activation, thrombosis, extravasation of white blood cells and, eventually, rejection. Pig antigens reacting with human, natural antibodies have turned out to be carbohydrates (7-10); the major one being the Galα1,3Gal epitope which is not expressed in old world monkeys, apes and humans due to an inactivation of the α1,3 galactosyltransferase (GT) (10-12).
Several methods have been proposed for the removal or elimination of xenoreactive antibodies from the blood of a recipient. Bach et al (Xenotransplantation, Eds: Cooper, D. K. C., et al, Springer Verlag, 1991, Chapter 6) proposed the perfusion of the recipients blood through an organ of the proposed donor species prior to transplantation of another, fresh organ, whereby anti-pig antibodies were removed.
Plasmapheresis has also been proposed for a non-specific removal of naturally occurring antibodies, whereby the graft survival is prolonged (e.g. Cairns et al, Rydberg et al). However, conventional plasmapheresis, or plasma exchange, results in loss of blood volume, which in turn may require a volume replacement with pooled preparations of fresh frozen plasma, human albumin, immunoglobulin etc. In addition, coagulation factors, platelets and antithrombotic factors must also be replaced. Such a treatment carries not only the risk of virus transfer, such as HIV, but also the risk of an anaphylactic reaction to foreign substances. Other negative side effects of plasmapheresis are recipient sensitization and activation of the complement and clotting system. Accordingly, plasmapheresis does not appear to be either practical or safe.
Other methods for the removal of xenoreactive antibodies involve non-specific antibody removal. Protein A, a major component of the cell wall of S. aureus, has a high affinity for a portion of the Fc-region of sub-classes 1, 2 and 4 of immunoglobulin G (IgG1, IgG2, IgG4) and has been used for the non-specific removal of anti-HLA antibodies from hypersensitized patients in need of kidney transplants. The efficacy of the Protein A column treatment after kidney transplantation have been reported (Dantal J., et al, New England J. Med. 550: 7-14, 1994; Nilsson, I. M. et al, Blood 58: 38-44, 1981; Palmer, A., et al., The Lancet Jan. 7, 1989, pp. 10-12). One essential drawback with the use of a Protein A column technique in the context of xenotransplantation is, however, the fact that only IgG will be removed. Lately, it has been shown that the antibodies involved in HAR during a transplantation from pig to human may involve several other immunoglobulin classes. In addition, the non-specific antibody removal will cause a general deterioration of the patients immune defense, which quite naturally is not desirable during such a process as a transplantation procedure, where the patient is immunosuppressed.
Leventhal et al (WO 95/31209) propose a method of preventing or ameliorating a hyperacute reaction occurring after transplantation of a pig organ to a primate recipient, including a human. The method involves passing the recipients plasma over a column with a coupled protein, which binds to and thereby removes immunoglobulin therefrom. The protein is selected from a group consisting of Staphylococcus aureus protein A, Streptococcus protein G and anti-human immunoglobulin antibodies. This method suffers from the same drawbacks as depicted above for the protein A column.
It has been shown (Platt et al, Good et al, Holgersson et al) that pig antigens reacting with human, natural antibodies are carbohydrates, the major one being the Galα1,3Gal epitope, which is not expressed in old world monkeys, apes and humans due to an inactivation of the α1,3 galactosyltransferase.
Recently, McKenzie et al showed that COS cells transfected with the α1,3-galactosyltransferase cDNA expressed the Galα1,3Gal-epitope on their surfaces and could absorb most of the human anti-pig activity from human serum.
Further, Galα1,3Gal-derivatized columns have been used to specifically remove anti-pig activity from human serum (22), free Galα1,3Gal disaccharides have been shown to prevent binding of anti-pig antibodies to porcine cells, including endothelium (23), and so has porcine stomach mucin (24). However, organic synthesis of saccharides is a very laborious and expensive method, which in addition is rather slow, and accordingly, has not found any wide spread applicability.
Apart from the HAR, xenografts are still typically rejected within days in a process that has been termed delayed xenograft rejection (DXR) (29). DXR is characterized by mononuclear cell activation and graft infiltration, as well as cytokine production (29). The importance of the Galα1,3Gal epitope for these cellular events is not known, although human anti-Galα1,3Gal antibodies were recently shown to be involved in antibody-dependent cellular cytotoxicity (ADCC) of porcine cells (30).
Thus, there is still a need of cheaper and more efficient methods for the elimination of foreign antibodies, such as pig-antibodies, from the blood from a recipient who is to obtain a xenotransplant. In addition, within the field of xenotransplantation, of methods for the prevention of DXR.